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- EMDB-19340: SPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local r... -

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Basic information

Entry
Database: EMDB / ID: EMD-19340
TitleSPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local refinement
Map dataEMhancer: SPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local refinement
Sample
  • Complex: Local refinement of GFP part of the SPNS2:sfGFP heterodimer assembled by Di-Gluebody
    • Complex: Green fluorescent protein
    • Complex: Di-Gluebody
    • Protein or peptide: Green fluorescent protein
    • Protein or peptide: Gluebody GbEnhancer
    • Protein or peptide: Gluebody GbC4
KeywordsDNA helicase / Di-Gluebody / HYDROLASE
Function / homologyGreen fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / bioluminescence / generation of precursor metabolites and energy / Green fluorescent protein
Function and homology information
Biological speciesAequorea victoria (jellyfish) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.75 Å
AuthorsYi G / Ye M / Mamalis D / Sauer DB / von Delft F / Davis BG / Gilbert RJC
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Other private United Kingdom
CitationJournal: To Be Published
Title: Di-Gluebodies: Rigid modular nanobody protein assemblies enabling simultaneous determination of high-resolution cryo-EM structures
Authors: Yi G / Mamalis D / Ye M / Carrique L / Fairhead M / Li H / Duerr K / Zhang P / Sauer DB / von Delft F / Davis BG / Gilbert RJC
History
DepositionJan 2, 2024-
Header (metadata) releaseJan 15, 2025-
Map releaseJan 15, 2025-
UpdateJan 15, 2025-
Current statusJan 15, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19340.png

Downloads & links

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Map

FileDownload / File: emd_19340.map.gz / Format: CCP4 / Size: 163.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEMhancer: SPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local refinement
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.93 Å/pix.
x 350 pix.
= 326.2 Å		0.93 Å/pix.
x 350 pix.
= 326.2 Å		0.93 Å/pix.
x 350 pix.
= 326.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.932 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-0.00482989 - 2.2700202
Average (Standard dev.)0.0015709491 (±0.024552234)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions350350350
Spacing350350350
CellA=B=C: 326.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19340_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: SPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP...

Fileemd_19340_additional_1.map
AnnotationSPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local refinement
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map A

Fileemd_19340_half_map_1.map
Annotationhalf map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_19340_half_map_2.map
Annotationhalf map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Local refinement of GFP part of the SPNS2:sfGFP heterodimer assem...

EntireName: Local refinement of GFP part of the SPNS2:sfGFP heterodimer assembled by Di-Gluebody
Components
  • Complex: Local refinement of GFP part of the SPNS2:sfGFP heterodimer assembled by Di-Gluebody
    • Complex: Green fluorescent protein
    • Complex: Di-Gluebody
    • Protein or peptide: Green fluorescent protein
    • Protein or peptide: Gluebody GbEnhancer
    • Protein or peptide: Gluebody GbC4

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Supramolecule #1: Local refinement of GFP part of the SPNS2:sfGFP heterodimer assem...

SupramoleculeName: Local refinement of GFP part of the SPNS2:sfGFP heterodimer assembled by Di-Gluebody
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: GbEnhancer and GbC4 were assembled via a disulfide to form a hetero Di-Gluebody
Molecular weightTheoretical: 112 KDa

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Supramolecule #2: Green fluorescent protein

SupramoleculeName: Green fluorescent protein / type: complex / ID: 2 / Parent: 1 / Details: Green fluorescent protein
Source (natural)Organism: Aequorea victoria (jellyfish)

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Supramolecule #3: Di-Gluebody

SupramoleculeName: Di-Gluebody / type: complex / ID: 3 / Parent: 1 / Details: Gluebody GbEnhancer and Gluebody GbC4
Source (natural)Organism: Lama glama (llama)

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Macromolecule #1: Green fluorescent protein

MacromoleculeName: Green fluorescent protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Aequorea victoria (jellyfish)
Molecular weightTheoretical: 26.81923 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSKGEELFTG VVPILVELDG DVNGHKFSVR GEGEGDATNG KLTLKFICTT GKLPVPWPTL VTTLTYGVQC FSRYPDHMKR HDFFKSAMP EGYVQERTIS FKDDGTYKTR AEVKFEGDTL VNRIELKGID FKEDGNILGH KLEYNFNSHN VYITADKQKN G IKANFKIR ...String:
MSKGEELFTG VVPILVELDG DVNGHKFSVR GEGEGDATNG KLTLKFICTT GKLPVPWPTL VTTLTYGVQC FSRYPDHMKR HDFFKSAMP EGYVQERTIS FKDDGTYKTR AEVKFEGDTL VNRIELKGID FKEDGNILGH KLEYNFNSHN VYITADKQKN G IKANFKIR HNVEDGSVQL ADHYQQNTPI GDGPVLLPDN HYLSTQSVLS KDPNEKRDHM VLLEFVTAAG ITHGMDELYK

UniProtKB: Green fluorescent protein

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Macromolecule #2: Gluebody GbEnhancer

MacromoleculeName: Gluebody GbEnhancer / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 12.689139 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
QVQLVENGGA CVKPGGSLRL SCAASGFPVN RYSMRWYRQA PGKEREWVAG MSSAGDRSSY EDSVKGRFTI SRDDARNTVY LQMNSLKPE DTAVYYCNVN VGFEYWGQGT QVMVS

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Macromolecule #3: Gluebody GbC4

MacromoleculeName: Gluebody GbC4 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 12.982609 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SQGQLVENGG GCVKAGGSLR LSCAASQGTL SNLVTGWFRR APGKEREFVA NIGRDGLTVY SNSVKGRFTI SRDRAKNTVY LQMDSLKPE DTAVYYCAGR LSRFPGEYDY WSKGTPVMVS

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration4.2 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
150.0 mMNaClsodium chloride
0.025 %C24H46O11DDM
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number real images: 8674 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1127724
Startup modelType of model: INSILICO MODEL / In silico model: Ab-initio reconstruction
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.75 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0) / Number images used: 321178
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0) / Details: Non-uniform refinement in cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0) / Details: Local refinement in cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

3k1k


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Overall B value: 154.9
Output model

PDB-8rle:
SPNS2:sfGFP hetero dimer assembled by Di-Gluebody - sfGFP local refinement

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